Sex-dependent vulnerability of fetal nonhuman primate cardiac mitochondria to moderate maternal nutrient reduction.

Poor maternal nutrition in pregnancy affects fetal development, predisposing offspring to cardiometabolic diseases. The role of mitochondria during fetal development on later-life cardiac dysfunction caused by maternal nutrient reduction (MNR) remains unexplored. We hypothesized that MNR during gestation causes fetal cardiac bioenergetic deficits, compromising cardiac mitochondrial metabolism and reserve capacity. To enable human translation, we developed a primate baboon model (Papio spp.) of moderate MNR in which mothers receive 70% of control nutrition during pregnancy, resulting in intrauterine growth restriction (IUGR) offspring and later exhibiting myocardial remodeling and heart failure at human equivalent ∼25 years. Term control and MNR baboon offspring were necropsied following cesarean-section, and left ventricle (LV) samples were collected. MNR adversely impacted fetal cardiac LV mitochondria in a sex-dependent fashion. Increased maternal plasma aspartate aminotransferase, creatine phosphokinase (CPK), and elevated cortisol levels in MNR concomitant with decreased blood insulin in male fetal MNR were measured. MNR resulted in a two-fold increase in fetal LV mitochondrial DNA (mtDNA). MNR resulted in increased transcripts for several respiratory chain (NDUFB8, UQCRC1, and cytochrome c) and adenosine triphosphate (ATP) synthase proteins. However, MNR fetal LV mitochondrial complex I and complex II/III activities were significantly decreased, possibly contributing to the 73% decreased ATP content and increased lipid peroxidation. MNR fetal LV showed mitochondria with sparse and disarranged cristae dysmorphology. Conclusion: MNR disruption of fetal cardiac mitochondrial fitness likely contributes to the documented developmental programming of adult cardiac dysfunction, indicating a programmed mitochondrial inability to deliver sufficient energy to cardiac tissues as a chronic mechanism for later-life heart failure.

Liraglutide Protects Against Brain Amyloid-ß1-42 Accumulation in Female Mice with Early Alzheimer's Disease-Like Pathology by Partially Rescuing Oxidative/Nitrosative Stress and Inflammation.

Alzheimer's disease (AD) is the most common form of dementia worldwide, being characterized by the deposition of senile plaques, neurofibrillary tangles (enriched in the amyloid beta (Aß) peptide and hyperphosphorylated tau (p-tau), respectively) and memory loss. Aging, type 2 diabetes (T2D) and female sex (especially after menopause) are risk factors for AD, but their crosslinking mechanisms remain unclear. Most clinical trials targeting AD neuropathology failed and it remains incurable. However, evidence suggests that effective anti-T2D drugs, such as the GLP-1 mimetic and neuroprotector liraglutide, can be also efficient against AD. Thus, we aimed to study the benefits of a peripheral liraglutide treatment in AD female mice. We used blood and brain cortical lysates from 10-month-old 3xTg-AD female mice, treated for 28 days with liraglutide (0.2 mg/kg, once/day) to evaluate parameters affected in AD (e.g., Aß and p-tau, motor and cognitive function, glucose metabolism, inflammation and oxidative/nitrosative stress). Despite the limited signs of cognitive changes in mature female mice, liraglutide only reduced their cortical Aß1-42 levels. Liraglutide partially attenuated brain estradiol and GLP-1 and activated PKA levels, oxidative/nitrosative stress and inflammation in these AD female mice. Our results support the earlier use of liraglutide as a potential preventive/therapeutic agent against the accumulation of the first neuropathological features of AD in females.

Alzheimer's disease and type 2 diabetes-related alterations in brain mitochondria, autophagy and synaptic markers.

We aimed to investigate mitochondrial function, biogenesis and autophagy in the brain of type 2 diabetes (T2D) and Alzheimer's disease (AD) mice. Isolated brain mitochondria and homogenates from cerebral cortex and hippocampus of wild-type (WT), triple transgenic AD (3xTg-AD) and T2D mice were used to evaluate mitochondrial functional parameters and protein levels of mitochondrial biogenesis, autophagy and synaptic integrity markers, respectively. A significant decrease in mitochondrial respiration, membrane potential and energy levels was observed in T2D and 3xTg-AD mice. Also, a significant decrease in the levels of autophagy-related protein 7 (ATG7) and glycosylated lysosomal membrane protein 1 (LAMP1) was observed in cerebral cortex and hippocampus of T2D and 3xTg-AD mice. Moreover, both brain regions of 3xTg-AD mice present lower levels of nuclear respiratory factor (NRF) 1 while the levels of NRF2 are lower in both brain regions of T2D and 3xTg-AD mice. A decrease in mitochondrial encoded, nicotinamide adenine dinucleotide dehydrogenase subunit 1 (ND1) was also observed in T2D and 3xTg-AD mice although only statistically significant in T2D cortex. Furthermore, a decrease in the levels of postsynaptic density protein 95 (PSD95) in the cerebral cortex of 3xTg-AD mice and in hippocampus of T2D and 3xTg-AD mice and a decrease in the levels of synaptosomal-associated protein 25 (SNAP 25) in the hippocampus of T2D and 3xTg-AD mice were observed suggesting synaptic integrity loss. These results support the idea that alterations in mitochondrial function, biogenesis and autophagy cause synaptic damage in AD and T2D.

Clinical and Epidemiological Aspects of Scorpionism in the World: A Systematic Review.

OBJECTIVE: Scorpion stings are registered worldwide, but the incidence and the features of the envenomations vary depending on the region. The aim of this review was to summarize the epidemiological, clinical, diagnostic, and therapeutic data worldwide regarding humans stung by scorpions. METHODS: A systematic review of the literature was conducted through the online databases of the Virtual Health Library (VHL), which hosts Medline and the Latin American and Caribbean Center on Health Sciences Informational (LILACS) database. We selected articles published between January 1, 2002 and July 31, 2014. RESULTS: Scorpion envenomation reports were found throughout the world, mainly in subtropical and tropical regions. The clinical manifestations were sympathetically and parasympathetically mediated, depending on the species of scorpion. Some of the most common severe complications of scorpionism included respiratory distress syndrome, pulmonary edema, cardiac dysfunction, impaired hemostasis, pancreatitis, and multiple organ failure. Scorpion envenomation could be classified as mild, moderate, and severe, and the therapeutic approach was based on the case severity. The treatment comprised 3 components: symptomatic measures, vital functions support, and injection of antivenom. Moreover, the time that elapsed between the sting and administration of the appropriate medical care was extremely important to the patient's prognosis. CONCLUSIONS: The large number of scorpion stings worldwide is concerning and reaffirms the need for new prevention measures and policies to reduce the incidence, prevalence, morbidity, and mortality rates from these poisonous arachnids.

Pre-diabetes alters testicular PGC1-α/SIRT3 axis modulating mitochondrial bioenergetics and oxidative stress.

Pre-diabetes, a risk factor for type 2 diabetes development, leads to metabolic changes at testicular level. Peroxisome proliferator-activated receptor γ coactivator 1 α (PGC-1α) and Sirtuin 3 (Sirt3) are pivotal in mitochondrial function. We hypothesized that pre-diabetes disrupts testicular PGC-1α/Sirt3 axis, compromising testicular mitochondrial function. Using a high-energy-diet induced pre-diabetic rat model, we evaluated testicular levels of PGC-1α and its downstream targets, nuclear respiratory factors 1 (NRF-1) and 2 (NRF-2), mitochondrial transcription factor A (TFAM) and Sirt3. We also assessed mitochondrial DNA (mtDNA) content, mitochondrial function, energy levels and oxidative stress parameters. Protein levels were quantified by Western Blot, mtDNA content was determined by qPCR. Mitochondrial complex activity and oxidative stress parameters were spectrophotometrically evaluated. Adenine nucleotide levels, adenosine and its metabolites (inosine and hypoxanthine) were determined by reverse-phase HPLC. Pre-diabetic rats showed increased blood glucose levels and impaired glucose tolerance. Both testicular PGC-1α and Sirt3 levels were decreased. NRF-1, NRF-2 and TFAM were not altered. Testicular mtDNA content was decreased. Mitochondrial complex I activity was increased, whereas mitochondrial complex III activity was decreased. Adenylate energy charge was decreased in pre-diabetic rats, as were ATP and ADP levels. Conversely, AMP levels were increased, evidencing a decreased ATP/AMP ratio. Concerning to oxidative stress pre-diabetes decreased testicular antioxidant capacity and increased lipid and protein oxidation. In sum, pre-diabetes compromises testicular mitochondrial function by repressing PGC-1α/Sirt3 axis and mtDNA copy number, declining respiratory capacity and increasing oxidative stress. This study gives new insights into overall testicular bioenergetics at this prodromal stage of disease.

Effects of methylglyoxal and pyridoxamine in rat brain mitochondria bioenergetics and oxidative status.

Advanced glycation end products (AGEs) and methylglyoxal (MG), an important intermediate in AGEs synthesis, are thought to contribute to protein aging and to the pathogenesis of age-and diabetes-associated complications. This study was intended to investigate brain mitochondria bioenergetics and oxidative status of rats previously exposed to chronic treatment with MG and/or with pyridoxamine (PM), a glycation inhibitor. Brain mitochondrial fractions were obtained and several parameters were analyzed: respiratory chain [states 3 and 4 of respiration, respiratory control ratio (RCR), and ADP/O index] and phosphorylation system [transmembrane potential (ΔΨm), ADP-induced depolarization, repolarization lag phase, and ATP levels]; hydrogen peroxide (H2O2) production levels, mitochondrial aconitase activity, and malondialdehyde levels as well as non-enzymatic antioxidant defenses (vitamin E and glutathione levels) and enzymatic antioxidant defenses (glutathione disulfide reductase (GR), glutathione peroxidase (GPx), and manganese superoxide dismutase (MnSOD) activities). MG treatment induced a statistical significant decrease in RCR, aconitase and GR activities, and an increase in H2O2 production levels. The administration of PM did not counteract MG-induced effects and caused a significant decrease in ΔΨm. In mitochondria from control animals, PM caused an adaptive mechanism characterized by a decrease in aconitase and GR activities as well as an increase in both α-tocopherol levels and GPx and MnSOD activities. Altogether our results show that high levels of MG promote brain mitochondrial impairment and PM is not able to reverse MG-induced effects.

Insulin-induced recurrent hypoglycemia exacerbates diabetic brain mitochondrial dysfunction and oxidative imbalance.

Intensive insulin therapy can prevent or slow the progression of long-term diabetes complications but, at the same time, it increases the risk for episodes of severe hypoglycemia. In our study, we used a protocol intended to mimic the levels of blood glucose that occur in type 1 diabetic patients under an intensive insulin therapy. Streptozotocin (STZ)-induced diabetic rats were treated subcutaneously with twice-daily insulin injections for 2weeks to induce hypoglycemic episodes. Brain cortical and hippocampal mitochondria were isolated and mitochondrial bioenergetics (respiratory chain and phosphorylation system) and oxidative status parameters (malondialdehyde (MDA) levels, mitochondrial aconitase activity and enzymatic and non-enzymatic antioxidant defenses) were analyzed. The protein levels of synaptophysin, a marker of synaptic integrity, and caspase 9 activity were also evaluated in cortical and hippocampal homogenates. Brain cortical mitochondria isolated from hyper- and recurrent hypoglycemic animals presented higher levels of MDA and α-tocopherol together with an increased glutathione disulfide reductase activity, lower manganese superoxide dismutase (MnSOD) activity and glutathione-to-glutathione disulfide (GSH/GSSG) ratio. No significant alterations were found in cortical mitochondrial respiratory chain and oxidative phosphorylation system. Hippocampal mitochondria from both experimental groups presented an impaired oxidative phosphorylation system characterized by a decreased mitochondrial energization potential and ATP levels and higher repolarization lag phase. In addition, higher MDA levels and decreased GSH/GSSG, α-tocopherol levels, and aconitase, glutathione peroxidase and MnSOD activities were observed in both groups of animals. Hippocampal mitochondria from recurrent hypoglycemic animals also showed an impairment of the respiratory chain characterized by a lower state 3 of respiration, respiratory control ratio and ADP/O index, and a higher state 4 of respiration. Additionally, a non-statistically significant decrease in synaptophysin protein levels was observed in cortical homogenates from recurrent hypoglycemic rats as well as in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic rats. An increase in caspase 9 activity was also observed in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic animals. Our results show that mitochondrial dysfunction induced by long-term hyperglycemic effects is exacerbated by recurrent hypoglycemia, which may compromise the function and integrity of brain cells.

Mitochondrial dysfunction is the focus of quaternary ammonium surfactant toxicity to mammalian epithelial cells.

Surfactants have long been known to have microbicidal action and have been extensively used as antiseptics and disinfectants for a variety of general hygiene and clinical purposes. Among surfactants, quaternary ammonium compounds (QAC) are known to be the most useful antiseptics and disinfectants. However, our previous toxicological studies showed that QAC are also the most toxic surfactants for mammalian cells. An understanding of the mechanisms that underlie QAC toxicity is a crucial first step in their rational use and in the design and development of more effective and safer molecules. We show that QAC-induced toxicity is mediated primarily through mitochondrial dysfunction in mammalian columnar epithelial cell cultures in vitro. Toxic effects begin at sublethal concentrations and are characterized by mitochondrial fragmentation accompanied by decreased cellular energy charge. At very low concentrations, several QAC act on mitochondrial bioenergetics through a common mechanism of action, primarily by inhibiting mitochondrial respiration initiated at complex I and, to a lesser extent, by slowing down coupled ADP phosphorylation. The result is a reduction of cellular energy charge which, when reduced below 50% of its original value, induces apoptosis. The lethal effects are shown to be primarily a result of this process. At higher doses (closer to the critical micellar concentration), QAC induce the complete breakdown of cellular energy charge and necrotic cell death.

UCP2 and ANT differently modulate proton-leak in brain mitochondria of long-term hyperglycemic and recurrent hypoglycemic rats.

A growing body of evidence suggests that mitochondrial proton-leak functions as a regulator of reactive oxygen species production and its modulation may limit oxidative injury to tissues. The main purpose of this work was to characterize the proton-leak of brain cortical mitochondria from long-term hyperglycemic and insulin-induced recurrent hypoglycemic rats through the modulation of the uncoupling protein 2 (UCP2) and adenine nucleotide translocator (ANT). Streptozotocin-induced diabetic rats were treated subcutaneously with twice-daily insulin injections during 2 weeks to induce the hypoglycemic episodes. No differences in the basal proton-leak, UCP2 and ANT protein levels were observed between the experimental groups. Mitochondria from recurrent hypoglycemic rats presented a decrease in proton-leak in the presence of GDP, a specific UCP2 inhibitor, while an increase in proton-leak was observed in the presence of linoleic acid, a proton-leak activator, this effect being reverted by the simultaneous addition of GDP. Mitochondria from long-term hyperglycemic rats showed an enhanced susceptibility to ANT modulation as demonstrated by the complete inhibition of basal and linoleic acid-induced proton-leak caused by the ANT specific inhibitor carboxyatractyloside. Our results show that recurrent-hypoglycemia renders mitochondria more susceptible to UCPs modulation while the proton-leak of long-term hyperglycemic rats is mainly modulated by ANT, which suggest that brain cortical mitochondria have distinct adaptation mechanisms in face of different metabolic insults.

Mechanism of inhibition of mitochondrial ATP synthase by 17ß-estradiol.

17ß-estradiol (E2) is considered to modulate the ATP synthase activity through direct binding to the oligomycin sensitive-conferring protein. We have previously demonstrated that E2 increases the amplitude of depolarization associated with the addition of ADP to energized mitochondria (i.e., to initiate a phosphorylative cycle) suggesting a direct action on the phosphorylative system of mitochondria. The purpose of the present study was to investigate the underlying mechanisms responsible for this effect. We show here that E2 modulates the activity of mitochondrial ATP synthase by promoting the intrinsic uncoupling ("slipping") of the ATP synthase. E2 depressed RCR, ADP/O ratio and state 3 respiration, whereas state 4 respiration was increased and VFCCP (uncoupled respiration) remained unaltered. In contrast to the stimulatory effect on state 4 respiration, state 2 respiration and Volig were not affected by E2. The effect of E2 appeared to be directed towards ATP synthase, since glutamate/malate respiration, uncoupled from the electron transport chain, was unaffected by E2. Apparently, E2 allows a proton back-leak through the Fo component of ATP synthase. This action of E2 is dependent on the presence of ATP, is more pronounced at high membrane potentials, and it is reversed by oligomycin (a Fo-ATP synthase inhibitor) but not by resveratrol (a F1-ATP synthase inhibitor). Altogether, our data provide a mechanistic explanation for the effect of E2 at the level of mitochondrial ATP synthase.

The antiestrogen 4-hydroxytamoxifen protects against isotretinoin-induced permeability transition and bioenergetic dysfunction of liver mitochondria: comparison with tamoxifen.

The combination of isotretinoin (13-cis-retinoic acid) with antiestrogens seems to be a promising strategy for cancer chemotherapy. The aim of the study was to evaluate the effects of isotretinoin alone or in combination with 4-hydroxytamoxifen (OHTAM) and with its prodrug tamoxifen (TAM), on the functions of rat liver mitochondria, i.e., mitochondrial permeability transition (MPT), bioenergetic functions and adenine nucleotide translocase (ANT). Isotretinoin (5 nmol/mg protein) induced the Ca²âº-dependent MPT pore opening in mitochondria energized with succinate, which was prevented by OHTAM, cyclosporine A, TAM and ANT ligands. When mitochondria were energized with glutamate/malate and in the absence of added Ca²âº isotretinoin decreased the state 3 respiration, the ATP levels, the active ANT content and increased the lag phase of the phosphorylation cycle, demonstrating that isotretinoin decreased the mitochondrial phosphorylation efficiency. These changes of isotretinoin in bioenergetic parameters were not significant in the presence of succinate. The effects of isotretinoin at 5 nmol/mg protein on the Ca²âº-dependent MPT and phosphorylative efficacy may be related with interactions with the ANT. Above 10 nmol/mg protein isotretinoin strongly diminished the active ANT content, decreased the Δψ, inhibited the complex I and induced proton leak through the Fo fraction of complex V. The combination of OHTAM with isotretinoin only induced significant changes in the energy production systems at concentrations ≥5 nmol isotretinoin/mg protein. Therefore, our results suggest that isotretinoin-associated liver toxicity is possibly related with mitochondrial dysfunctions and that the combination with OHTAM may contribute to decrease its toxicity.

The antiestrogen endoxifen protects rat liver mitochondria from permeability transition pore opening and oxidative stress at concentrations that do not affect the phosphorylation efficiency.

Endoxifen (EDX) is a key active metabolite of tamoxifen (TAM) with higher affinity and specificity to estrogen receptors that also inhibits aromatase activity. It is safe and well tolerated by healthy humans, but its use requires toxicological characterization. In this study, the effects of EDX on mitochondria, the primary targets for xenobiotic-induced toxicity, were monitored to clarify its potential side effects. EDX up to 30 nmol/mg protein did not affect the mitochondrial oxidative phosphorylation. At 50 nmol EDX/mg protein, EDX decreased the ADP phosphorylation rate and a partial collapse of mitochondrial membrane potential (Δψ), that parallels a state 4 stimulation, was observed. As the stimulation of state 4 was not inhibited by oligomycin and 50 nmol EDX/mg protein caused a slight decrease in the light scattering of mitochondria, these data suggest that EDX promotes membrane permeabilization to protons, whereas TAM at the same concentration induced mitochondrial membrane disruption. Moreover, EDX at 10 nmol/mg protein prevented and reversed the Ca(2+)-induced depolarization of ΔΨ and the release of mitochondrial Ca(2+), similarly to cyclosporine A, indicating that EDX did not affect Ca(2+) uptake, but directly interfered with the proteins of the mitochondrial permeability transition (MPT) megacomplex, inhibiting MPT induction. At this concentration, EDX exhibited antioxidant activity that may account for the protective effect against MPT pore opening. In conclusion, EDX within the range of concentrations reached in tissues did not significantly damage the bioenergetic functions of mitochondria, contrarily to the prodrug TAM, and prevented the MPT pore opening and the oxidative stress in mitochondria, supporting that EDX may be a less toxic drug for women with breast carcinoma.

Statin restores cardiac autonomic response to acute hypoxia in hypercholesterolaemia.

BACKGROUND: Hypercholesterolaemia may alter cardiovascular autonomic function. We investigated the autonomic cardiovascular regulation during normoxia and hypoxia in familial isolated HC patients with or without statin treatment. MATERIALS AND METHODS: Low (LF-RR) and high (HF-RR) components of spectral analysis of RR interval and systolic arterial pressure (LF-SAP) were obtained during 5 min of normoxia and isocapnic hypoxia (10% O(2) ) in 10 normotensive familial HC patients without medication, in seven HC patients after a 12-week treatment period with 40 mg of simvastatin (HC + SVT) and in eight matched normal volunteers (CO). RESULTS: The HC patients had significant impairment of cardiac autonomic modulation parameters compared with CO at normoxia, which was maintained or even accentuated during hypoxia; these parameters included lower total variance of RR, increased normalized LF-RR, decreased normalized HF-RR, increased LF-RR/HF-RR ratio, higher LF-SAP component and reduced α index. However, the HC + SVT group had a significant improvement in all parameters: the LF-RR and LF-SAP decreased (indicating a decrease in cardiac and vascular sympathetic activity), the HF-RR increased (indicating an increase in parasympathetic activity) and the spontaneous baroreflex sensitivity improved. These changes were detected at normoxia and were maintained during hypoxia. CONCLUSIONS: Our data are the first to show that isolated HC is characterized by an increase in cardiac and vasomotor sympathetic drive, a decrease in cardiac vagal modulation and baroreflex impairment during normoxia and hypoxia. In addition, our data suggest that statin treatment has a potential role in restoring the physiological cardiovascular autonomic control at baseline and during cardiovascular challenge.

Cyanide preconditioning protects brain endothelial and NT2 neuron-like cells against glucotoxicity: role of mitochondrial reactive oxygen species and HIF-1α.

The current study was undertaken to address the role of mitochondrial reactive oxygen species (ROS), and hypoxia inducible factor-1 alpha (HIF-1α) signaling pathway in the protection against high glucose levels in brain endothelial and NT2 neuron-like cells. Rat brain endothelial cells (RBE4) treated with non-toxic concentrations of cyanide (≤1 µM; 1h) exhibited an increase in ROS levels, particularly hydrogen peroxide (H(2)O(2)). Cyanide also induced a modest mitochondrial depolarization, an increase in oxygen consumption and a structural (smaller mitochondria) and spatial (perinuclear region) reorganization of mitochondrial network. The stabilization and nuclear activation of HIF-1α in the presence of cyanide were also observed, which resulted in an increase in vascular endothelial growth factor (VEGF), endothelial nitric oxide synthase (eNOS) and erythropoietin (EPO) protein levels reflecting an adaptive response. Importantly, preconditioning induced by cyanide protected brain endothelial cells against high glucose-mediated damage by the prevention of apoptotic cell death. In mitochondrial DNA-depleted NT2 (NT2 ρ0) cells, cyanide (0.1 µM) was unable to stimulate ROS production and, consequently, protect against glucotoxicity. Conversely, in NT2 cells, the parental cells with functional mitochondria, cyanide significantly increased ROS levels protecting against high glucose-induced neuronal cell loss and activation of caspase-3. The free radical scavenger N-acetyl-L-cysteine and the specific HIF-1α inhibitor 2-methoxyestradiol completely abolished the protective effects of cyanide preconditioning. Altogether our results demonstrate that mitochondrial preconditioning induced by cyanide triggers a protective response mediated by mitochondrial ROS and HIF-1α activation and signaling, which render brain endothelial and neuronal cells resistant against glucotoxicity.

Cortical and hippocampal mitochondria bioenergetics and oxidative status during hyperglycemia and/or insulin-induced hypoglycemia.

This study was undertaken to evaluate the effects of streptozotocin (STZ)-induced hyperglycemia and insulin-induced hypoglycemia in cortical and hippocampal mitochondria bioenergetics and oxidative status. For that purpose we used, citrate (vehicle)-treated Wistar rats, STZ-treated rats [i.p., 50mg/kg body weight] and STZ-treated rats injected with insulin [s.c., dose adjusted to blood glucose levels] 1h prior to sacrifice to induce an acute episode of hypoglycemia. Several parameters were analyzed: respiratory chain, phosphorylation system, thiobarbituric acid reactive substances (TBARS) levels, hydrogen peroxide (H(2)O(2)) production rate, and non-enzymatic and enzymatic antioxidant defenses. Cortical mitochondria from insulin-induced hypoglycemic rats present a significant decrease in the ADP/O index, a significant increase in the repolarization lag phase and a decrease in GSH/GSSG ratio when compared with STZ and control mitochondria. Both STZ-induced diabetes and insulin-induced hypoglycemia promote a significant increase in TBARS levels and a decrease in glutathione disulfide reductase activity. Diabetic cortical mitochondria present a significant decrease in glutathione peroxidase (GPx) activity compared to control mitochondria. In turn, insulin-induced hypoglycemia induced a significant increase in GPx and manganese superoxide dismutase (MnSOD) activities. In hippocampal mitochondria, insulin-induced hypoglycemia increases the respiratory control ratio whereas both situations, hyper- and hypoglycemia, potentiate H(2)O(2) production and decrease the activity of MnSOD. These results suggest that the poor glycemic control that occurs in type 1 diabetic patients undergoing insulin therapy may have detrimental effects in brain areas involved in learning and memory.

Effects of rapamycin and TOR on aging and memory: implications for Alzheimer's disease.

Rapamycin is a macrolide immunosuppressant drug, originally used as an anti-fungal agent, which is widely used in transplantation medicine to prevent organ rejection. Target of rapamycin (TOR) is an evolutionarily conserved serine/threonine kinase with pleiotropic cellular functions, regulating processes such as growth and metabolism, cell survival, transcription and autophagy. TOR intervenes in two distinct enzymatic complexes with different functions, a rapamycin-sensitive complex TORC1 and a rapamycin-insensitive complex TORC2. Rapamycin has an inhibitory effect on TORC1 activity and it has been suggested to increase life span, an effect correlated with decreased protein biosynthesis and autophagy activation. In the CNS, rapamycin shows beneficial effects in neuronal survival and plasticity, thus contributing to memory improvement. In this review, evidence implying rapamycin and TOR in aging/life span extension and memory improvement will be discussed. Recent findings about the effects of rapamycin on Alzheimer's disease-associated neuropathology will be also discussed.

Impact of STZ-induced hyperglycemia and insulin-induced hypoglycemia in plasma amino acids and cortical synaptosomal neurotransmitters.

In this work, we evaluated the effects of streptozotocin (STZ)-induced hyperglycemia and an acute episode of insulin-induced hypoglycemia in plasma amino acids and cortical neurotransmitters. For that purpose, we used citrate (vehicle)-treated Wistar rats, STZ-treated rats [i.p., 50 mg/kg body weight], and STZ-treated rats injected with insulin [s.c., dose adjusted with blood glucose levels] 1 h prior to sacrifice to induce an acute episode of hypoglycemia. Plasma was collected for determination of amino acids levels. In addition, cortical synaptosomal preparations were obtained and the total levels of neurotransmitters, levels of aspartate, glutamate, taurine, and GABA released by the action of KCl, iodoacetic acid (IAA), ouabain, and veratridine, membrane potential and ATP levels were evaluated. Compared with control rats, plasma from hypoglycemic rats presented increased levels of aspartate, glutamate, glutamine, and taurine whereas GABA levels were decreased in STZ and hypoglycemic rats. Similarly, glutamate and taurine levels were increased in hypoglycemic synaptosomes while GABA decreased in hypoglycemic and STZ-diabetic synaptosomes. The depolarizing agent KCl promoted an increase in aspartate, glutamate, and taurine release from hypoglycemic synaptosomes. The highest release of neurotransmitters occurred in the presence of veratridine and ouabain, two other depolarizing agents, in all groups of experimental animals. However, a higher release of glutamate was observed in the diabetic and hypoglycemic synaptosomes. No alterations were observed in synaptosomal membrane potential and ATP levels. These results show that in the presence of a metabolic insult a higher release of excitatory amino acids occurs, which may underlay the neuronal injury observed in type 1 diabetic patients under insulin therapy.

Coconut Oil Supplementation Does Not Affect Blood Pressure Variability and Oxidative Stress: A Placebo-Controlled Clinical Study in Stage-1 Hypertensive Patients.

Exploring an alternative to improve the clinical management of hypertension, we tested the hypothesis that food supplementation with coconut oil (EVCO), alone or combined with aerobic exercise training, could exert an antihypertensive effect (primary outcome) in patients with stage 1 hypertension. Forty-five hypertensive volunteers of both genders participated in a placebo-controlled clinical trial. The volunteers were submitted to 24-hour ambulatory blood pressure monitoring, analysis of blood pressure variability (BPV), measurement of serum malondialdehyde (MDA) and nutritional assessment. Results indicate that EVCO consumption had no adverse effects. The supplementation did not increase the caloric intake compared with placebo, and the dietary constituents were similar between groups, except for the saturated fats, especially lauric acid. The analysis of blood pressure indicated absence of antihypertensive effect of EVCO alone or combined with physical training. Furthermore, no effects on blood pressure variability and oxidative stress were observed in the supplemented hypertensive patients. Thus, despite the results observed in pre-clinical studies, the current clinical study did not provide evidence to support the use of coconut oil as an adjuvant in the management of hypertension in humans.

Kaolinite/cashew gum bionanocomposite for doxazosin incorporation and its release.

Incorporation of drugs in clay minerals has been widely proposed for the controlled-release or increased solubility of drugs. In this context, a bionanocomposite based on kaolinite and cashew gum (Kln/Gum) was synthesized and characterized by X-ray diffraction (XRD), thermal analysis (TG/DTA), and Fourier transform infrared spectroscopy (FTIR). The bionanocomposite was applied to the incorporation and further release of doxazosin mesylate (DB). The influence of solution pH (1-3), adsorbent dose (20-50 mg), initial drug concentration (20.0-70.0 mg L-1), contact time (15-300 min), and temperature (25, 35, and 45 °C) were systematically evaluated. Equilibrium was reached around 60 min, with a maximum adsorption capacity of 31.5 ± 2.0 mg g-1 at a pH of 3.0 and 25 °C. Hydrogen bonding contributed to DB incorporation on the Kln/Gum. In addition, DB maximum amounts of 16.80 ± 0.58 and 77.00 ± 2.46% were released at pH values of 1.2 and 7.4, respectively. These results indicated that the Kln/Gum bionanocomposite is an effective and promising material for the incorporation/release of drugs with similar structures to DB.

Insulin neuroprotection against oxidative stress is mediated by Akt and GSK-3beta signaling pathways and changes in protein expression.

Previously we demonstrated that insulin protects against neuronal oxidative stress by restoring antioxidants and energy metabolism. In this study, we analysed how insulin influences insulin-(IR) and insulin growth factor-1 receptor (IGF-1R) intracellular signaling pathways after oxidative stress caused by ascorbate/Fe2+ in rat cortical neurons. Insulin prevented oxidative stress-induced decrease in tyrosine phosphorylation of IR and IGF-1R and Akt inactivation. Insulin also decreased the active form of glycogen synthase kinase-3beta (GSK-3beta) upon oxidation. Since phosphatidylinositol 3-kinase (PI-3K)/Akt-mediated inhibition of GSK-3beta may stimulate protein synthesis and decrease apoptosis, we analysed mRNA and protein expression of "candidate" proteins involved in antioxidant defense, glucose metabolism and apoptosis. Insulin prevented oxidative stress-induced increase in glutathione peroxidase-1 and decrease in hexokinase-II expression, supporting previous findings of changes in glutathione redox cycle and glycolysis. Moreover, insulin precluded Bcl-2 decrease and caspase-3 increased expression. Concordantly, insulin abolished caspase-3 activity and DNA fragmentation caused by oxidative stress. Thus, insulin-mediated activation of IR/IGF-1R stimulates PI-3K/Akt and inhibits GSK-3beta signaling pathways, modifying neuronal antioxidant defense-, glucose metabolism- and anti-apoptotic-associated protein synthesis. These and previous data implicate insulin as a promising neuroprotective agent against oxidative stress associated with neurodegenerative diseases.